Long-Range Single-Molecule Förster Resonance Energy Transfer between Alexa Dyes in Zero-Mode Waveguides
Förster resonance energy transfer (FRET) is widely used as a molecular ruler to monitor biomolecular conformations and interactions dynamics, but FRET is generally limited to distances below 10 nm.
Last year, we showed that zero-mode waveguides (ZMW) nanoapertures can enable single molecule FRET detection at spatial distances exceeding 10 nm with higher FRET efficiencies. However, this earlier work was limited to a specific Atto 550 – Atto 647N fluorescent dye pair, rising the issue that observations of FRET enhancement could be an artefact related to this specific set of fluorescent dyes.
In a recent ACS Omega paper, we use a markedly different set of fluorescent dyes (Alexa Fluor 546 and Alexa Fluor 647). Our new single molecule FRET data quantitatively demonstrate enhanced FRET efficiencies at large separations exceeding 10 nm confirming our earlier conclusions.
- The FRET enhancement inside a ZMW does not depend on the set of fluorescent dyes, validating the ZMW approach.
- Nanoapertures and nanophotonics are demonstrated to extend the spatial range of FRET to distances where dipole-dipole interactions would otherwise be too weak to produce detectable FRET signals.
Also available on ArXiv 2004.04513